Syphilis is a disease triggered through infection with Treponema pallidum. By virtue of development of an effective therapeutic agent (e.g., penicillin), the incidence of syphilis has decreased since the 1940s. However, the incidence has tended to increase in recent years. One characteristic feature of recent years' new-type syphilis patients resides in that many syphilis patients have a complication of HIV infection. Conceivable reasons for such high complication rate are that syphilis and HIV are sexually transmitted diseases, and that syphilis increases the risk of HIV infection. Under such circumstances, early detection and treatment of syphilis patients are required for preventing prevalence of infection with syphilis and HIV.
Whether or not a subject has contracted syphilis is determined through immunologically detecting an anti-Treponema pallidum antibody in blood. Meanwhile, there are a large number of surface antigens on the surface of a cell of Treponema pallidum, and syphilis is detected through an immunological assay based on antigen-antibody reaction between the surface antigens and an anti-Treponema pallidum antibody present in a sample. Known surface antigens present on the surface of a cell of Treponema pallidum mainly include antigens having molecular weights of 47 kDa, 42 kDa, 37 kDa, 17 kDa, and 15 kDa.
Currently, the surface antigen of Treponema pallidum cells employed in diagnosis of syphilis is produced through culturing Treponema pallidum cells in the testicles of rabbits, solubilizing and extracting the cells with a surfactant or the like, and purifying the target cells through various methods for removing impurities. Since the thus-prepared antigen derived from Treponema pallidum has high specificity to an anti-Treponema pallidum antibody, the antigen enables early detection of syphilis in patients. However, when the antigen is produced through the aforementioned antigen production method employing rabbits, a certain limitation is imposed on the yield of antigen, due to use of the testicles of rabbits as hosts. In addition, the state of growth of Treponema pallidum varies among the host rabbits, and difficulty is countered in consistent production of Treponema pallidum in a large amount. Notably, at present, direct artificial culturing of Treponema pallidum has never been attained.
In recent years, there has been proposed a method for producing a surface antigen of Treponema pallidum cells through a recombination technique. The gene encoding syphilis 47 kDa antigen has already been cloned, and the amino acid sequence formed of 415 amino acids has been determined (see, for example, Non-Patent Documents 1 and 3). The crystal structure and biological roles of the syphilis 47 kDa antigen have already been reported. It has been elucidated that the antigen is known to have four structural domains A, B, C, and D (see, for example, Non-Patent Documents 2 and 3).
Non-Patent Document 3 discloses that domain A is formed of amino acid 1 to amino acid 34 (A1 domain) and amino acid 157 to amino acid 207 (A2 domain), that domain B is formed of amino acid 35 to amino acid 156, that domain C is formed of amino acid 208 to amino acid 335, and that domain D is formed of amino acid 336 to amino acid 415, the positions of the amino acids being counted from the N-terminus (see Non-Patent Document 3, FIG. 1).
The aforementioned 47 kDa antigen has been reported to have an antibody recognition site formed of an amino acid sequence having antigen activity (see, for example, Non-Patent Document 4).
There has been disclosed an anti-Treponema pallidum antibody assay method employing the aforementioned antigen. In the disclosed method, syphilis 47 kDa antigen is produced through a recombination technique, and an anti-Treponema pallidum antibody is immunologically determined by use of the produced antigen (see Patent Document 1). A similar assay method employing a fusion protein between the N-terminus of 15 kDa or 17 kDa antigen and glutathione-S-transferase is also disclosed (see Patent Document 2).
In addition to antigen production through a recombination technique, there is also disclosed another anti-Treponema pallidum antibody assay method including synthesizing a peptide having 47 kDa antigen activity and employing the peptide as an antigen (see Patent Document 3).